DESCRIPTION: The principal focus of this project is multiple opioid receptor binding sites. The principal investigator has a long-established interest in identifying multiple subtypes of opioid receptors, using a variety of techniques including receptor binding and analgesia studies. However, when the mu opioid receptor was cloned, it was clear that only one type of receptor was coded by the mu receptor gene, and that potential multiple receptor subtypes must arise from different mechanisms. During the previous funding period, the principal investigator has explored the concept that multiple mu receptors arise from alternative splice variants of the mu receptor (MOR-1) gene. These concepts arose originally from antisense studies of opioid analgesia, in which different antisense oligonucleotides representing different exons of the MOR-1 gene produced different effects on analgesia. The studies were furthered by sequencing of the MOR-1 gene encoding a number of introns and exons. Although their physiological significance is not yet clear, the mRNA coding for at least 12 variants of this receptor have been isolated. The proposed studies will further explore these potential receptor subtypes. The first specific aim will provide a detailed characterization of mu receptor splice variants, including pharmacological and anatomical studies and identification of new clones. The second aim will perform similar experiments on the ORL-1 receptor, identifying at least one novel splice variant of this receptor. The third aim will explore the possible existence of MOR-1/ORL-1 heterodimers, based upon the preliminary finding that co-expression of both of these receptors into cell lines produces binding sites which are different from either site alone. The fourth aim will compare the binding properties of several peptide ligands for the OPRL-1 receptor.